Keratinocyte growth factor (KGF) is a growth factor specific for epithelial cells that was first identified in conditioned medium of a human embryonic lung fibroblast cell line [Rubin et al., Proc. Natl. Acad. Sci. USA 86:802-806 (1989)]. Expression of messenger RNA for KGF has been detected in several stromal fibroblast cell lines derived from epithelial tissues at various stages of development. The transcript for KGF was also evident in RNA extracted from normal adult kidney and organs of the gastrointestinal tract [Finch et al., Science 245:752-755 (1989)]. Evidence that KGF is secreted from fibroblasts in culture and is expressed in vivo in the dermis but not the epidermis indicates that KGF may be an important normal paracrine effector of keratinocyte proliferation. Studies have shown that KGF is as potent as epidermal growth factor (EGF) in stimulating the proliferation of primary or secondary human keratinocytes in tissue culture [Marchese et al., J. Cell. Phys. 144:326-332 (1990)]. KGF is produced by mesenchymal cells near the epithelium of many organs including the epidermis, oral and lower gastrointestinal epithelium, pancreas, liver, lung, urothelium, prostate epithelium and others [Finch et al, supra, Housley et al., J Clin Invest. 94:1764-77, (1994); Yi et al., Am J Path. 145:80-85, (1994); Pierce et al., J Exp. Med. 179831-40, (1994); Yi et al, J Urol. 154:1566-70, (1995); and Ulich et al., J Clin Invest. 93:1298-1306, (1994)].
The purification of KGF from conditioned medium of a human embryonic fibroblast cell line, as well as the partial amino acid sequencing of purified KGF, the cloning of the KGF gene, and the expression of the gene in bacterial cells to yield biologically active recombinant KGF are described in International Patent Publication WO 90/08771. This publication also discloses that KGF or KGF-like polypeptides are useful as wound healing agents for burn wounds or to stimulate transplanted corneal tissue.
Ex vivo and in vivo studies in normal adult animals have shown that KGF-1 (hereinafter “KGF”) produces changes in hair follicle morphogenesis, hepatocyte proliferation, and epithelial cell proliferation in the lung, breast, pancreas, stomach, small intestine, and large intestine [Panos et al., J. Clin. Invest. 92:969-977 (1993); Ulich et al., Am. J. Path. 144:862-868 (1994); Yi et al., Am. J. Path. 145:80-85 (1994); and Ulich et al., J. Clin. Invest. 93:1298-1306 (1994)]. The role of KGF in embryonic or neonatal development is currently under investigation; however, KGF has been documented to be an important mediator of seminal vesicle development in the newborn mouse [Alarid et al., Proc. Natl. Acad. Sci. USA 91:1074-1078 (1994)]. Additionally, mice overexpressing KGF in hepatocytes exhibit polycystic kidneys [Nguyen et al., Oncogene 12:2109-19, (1996)], while KGF overexpresion in lung using a surfactant promoter result in mice with pulmonary cystademonas [Simonet et al., Proc. Natl. Acad. Sci. USA 92:12461-65, (1995)], demonstrating the importance of KGF in normal renal and pulmonary development.
KGF has been demonstrated to increase re-epithelialization and increased thickness of the epithelium when recombinant KGF was topically applied to wounds surgically induced in the rabbit ear or in porcine skin [Pierce et al., J. Exp. Med. 179:831-840 (1994]); and Staiano-Coico et al., J. Exp. Med. 178:865-878 (1993)]. Bosch, et al., [J. Clin. Invest. 98:2683-2687 (1996)] reported that administration of keratinocyte growth factor will induce the proliferation of liver cells.
Typically, purified polypeptides are only marginally stable in an aqueous state and undergo chemical and physical degradation resulting in a loss of biological activity during processing and storage. Additionally, polypeptide compositions in aqueous solution undergo hydrolysis, such as deamidation and peptide bond cleavage. These effects represent a serious problem for therapeutically active polypeptides which are intended to be administered to humans within a defined dosage range based on biological activity.
Administration of purified keratinocyte growth factor remains a promising candidate to treat many diseases that affect the human population. However, the ability of the KGF to remain a stable pharmaceutical composition over time in a variety of storage conditions and then be effective for patients in vivo has not been addressed. Thus, there remains a need in the art to provide keratinocyte growth factor in stable formulations that are useful as therapeutic agents to treat the variety of diseases which benefit from KGF-mediated stimulation of epithelial cell growth.